Facile synthesis of new N-(aminocycloalkylene)amino acid compounds using chiral triflate esters with N-Boc-aminopyrrolidines and N-Boc-aminopiperidines

In this study, we prepared a series of new N-(aminocycloalkylene)amino acid derivatives for use in chiral building blocks. The method was based on the conversion of enantiopure α-hydroxy acid esters into the corresponding chiral triflate esters, which were displaced by a nucleophilic substitution SN2 reaction with aminopyrrolidine and aminopiperidine derivatives, and the inversion of the configuration to give methyl 2-[(Boc-amino)cycloamin-1-yl]alkanoates with good yield and high enantiomeric and diastereomeric purity. Synthesized 2-[(Boc-amino)piperidin-1-yl]propanoates combined with ethyl l-phenylalaninate gave new chiral N-Boc- and N-nosyl-dipeptides containing a piperidine moiety. The structures were elucidated by 1H-, 13C-, and 15N-NMR spectroscopy, high-resolution mass spectrometry, and X-ray crystallography analyses.


Introduction
N-(u-Aminoalkylene)amino acids have many interesting biological activities and play important roles in medicinal chemistry and drug discovery for the pharmaceutical industry. [1][2][3][4] In biochemistry, N-(u-aminoalkylene)amino acids, such as N-(2aminoethyl)glycine I, are used for the synthesis of peptide nucleic acids (PNAs) (Fig. 1). [5][6][7][8] N-(u-Aminoalkylene)amino acids can be synthesized in various ways: alkylation, reductive amination, and the Mitsunobu reaction. 9 In the alkylation reaction, a molecule of ethylenediamine or its mono N-protected derivative reacts with a molecule of an a-halocarboxylic acid to form an N-(u-aminoalkylene)amino acid molecule. Byt and Gilon reported a method for the alkylation synthesis of N-(u-aminoalkylene)amino acids via the reaction of alkylenediamine (NH 2 (CH 2 ) n NH 2 , n = 2, 3, 6) with a-haloacetic acid at 25°C for 48 h. 10 This experiment had a yield of 53-72%, but used a large excess of alkylenediamine. When optically active a-halogenocarboxylic acids were used as alkylating agents with ethylenediamine, a S N 2 nucleophilic substitution occurred with the consequent inversion of conguration.
It is widely known that the bromo derivative of a-haloacetic acid provides a higher yield during the nucleophilic substitution reaction because of its leaving group ability compared to that of the chloroacetic acid derivative. 9 Feagin et al. reported the preparation of benzyl 2-{[2-(Boc-amino)ethyl]amino}acetate by reacting N-Boc-ethylenediamine and ethyl bromoacetate in the presence of triethylamine in acetonitrile at 70-80°C for 100 min with a good yield of 72%. 11 Sugiyama et al. reported that chiral diamine, (S)-Cbz-HNCH 2 CH(CH 3 )NH 2 , reacted with ethyl bromoacetate and yielded the target chiral compound III in the presence of potassium carbonate in DCM at room temperature, but undesired N,N-dialkylated compounds were observed in the mixture as well. 12 Sherer and Brugger reported the details of the reaction of tert-butyl N-(azetidin-3-yl) carbamate with ethyl 2-bromopropanoate in the presence of triethylamine in DCM at room temperature for 13 h, a sequence which provided ethyl 2-[3-(N-Boc-amino)azetidin-l-y]) propanoate IV at a yield of 44%. 13 The compound IV block was used to prepare polycyclic Toll-like receptor (TLR) antagonists useful in the treatment of immune disorders. 14 The two most commonly used direct reductive amination methods are described below. The rst method utilizes hydride reducing agents, particularly sodium cyanoborohydride -NaBH 3 CN. 15 Manna et al. reported that the treatment of (2S)-2-(N-Boc-amino)propanal with ethyl glycinate hydrochloride in methanol gave the corresponding imine which, aer the addition of NaBH 3 CN and acetic acid, gave the nal chiral glycine ester derivative V with a good yield 68%. 16 The second method is catalytic hydrogenation with platinum, palladium, or nickel catalysts. 17 For example, N-(2-aminoethyl)glycine (I) was prepared by the reaction of diaminoethane with glyoxylic acid monohydrate and over Pd/C in ethanol under hydrogen gas at atm. pressure and room temperature. 18 Dueholm and coworkers reported that the treatment of N-Boc-aminoaldehyde with methylglycinate hydrochloride in a solvent containing KOAc and a catalyst Pd/C under a hydrogen atmosphere afforded methyl-N-(2-Boc-aminoethyl)glycinate (71%). 19 The difficulty of obtaining amino aldehydes as compounds has been noted, as has their instability. 20 Falkevich et al. reported on the development and synthesis of N-(u-aminoalkylene)amino acid derivatives from N-Boc-bamino alcohols with N-o-nitrobenzenesulfonyl-protected (o-NBS-protected) amino acid esters using the Mitsunobu reaction. 21 For example, the treatment of (2S)-2-(methylamino) propan-1-ol with o-NBS-Gly-OEt under Mitsunobu conditions followed, by deprotection with thiophenol, yielded the chiral glycine ester derivative V. 16 Sabu et al. used the Mitsunobu reaction for the prepared N-(2-aminoethyl)glycine derivative VI containing the diglyme moiety. 22 Herein, we report the design and preparation of methyl 2-[(N-Boc-amino)cycloaminyl]alkanoates from chiral triate esters with chiral 3-Boc-aminopyrrolidine, 3-Boc-aminopiperidine and achiral 4-Boc-aminopiperidine. Such amino acid derivatives offer valuable properties as isosteres, new conformationally restricted chiral amino acids, and building blocks that can be used as potentially biologically active substances and peptides. [23][24][25][26][27] Results and discussion The synthetic strategy of methyl 2-[(Boc-amino)cycloaminyl] alkanoates is outlined in Scheme 1. The starting (R)-and (S)-2hydroxy acid esters 1a-c used in this study are commercially available or were prepared from their acid form (see Experimental section). The synthetic sequence was started by transforming a-hydroxy carboxylates (R)-1a and (S)-1a into the triate esters, methyl(2R)-and (2S)-2-[(tri-uoromethanesulfonyl)oxy]propanoates (R)-2a and (S)-2a, using triuoromethanesulfonic anhydride and pyridine in DCM. 28 A triate group is an excellent leaving group used in nucleophilic substitution reactions and has been shown to be signicantly superior to other leaving groups in the Walden inversion, where the inversion of a stereogenic centre in a chiral molecule takes place. 29,30 It is known that the reaction of enantiopure a-halocarboxylic acid esters with amines is accompanied by extensive racemization of a-amino esters; with a-methanesulfonyloxy and a-toluenesulfonyloxy carboxylic acid derivatives, both racemization and elimination products are formed due to the drastic conditions. 31 Therefore, triate esters with primary and secondary amines are known to give N-substituted a-aminocarboxylates in the S N 2 reaction, resulting in good chemical as well as optical yields. Another advantage of triate esters is that they can be generated in situ and used subsequently without isolation. 32 According to Effenberger et al. ethyl (S)-2-hydroxypropionate was converted to a triate ester and then treatment of the triate ester with N-benzyl-N-methylamine in dichloromethane at 0-20°C supplied ethyl-N-benzyl-N-methyl-D-alaninate (yield, 96%); 33 Nilsson et al. reported that methyl(S)-3-(benzyloxy)-2-[(triuoromethanesulfonyl)oxy]propanoate with 1-methylpiperazine maintained at −40°C in toluene containing DIPEA produced methyl(R)-3-(benzyloxy)-2-(4-methylpiperazin-1-yl)propanoates (yield, 66%; ee 96%). 34 In our work, the reaction of chiral triate esters (R)-2a or (S)-2a with 4-Boc-aminopiperidine in the presence of TEA in DCM at −50°C led to the formation of the enantiomeric pure 2-[(Bocamino)piperidinyl]propanoates, (S)-3a in a 84% yield, and (R)-3a in a 74% yield. The structural assignment of compounds (S)-3a and (R)-3a was readily deduced via detailed spectral data analysis. The IR spectrum of (S)-3a contained characteristic absorption bands, such as 1728 (C]O, ester) and 1681 (C]O, Boc) cm −1 . The 1 H NMR spectrum of compound (S)-3a revealed a characteristic resonance for the protons of the COOCH 3 group, which appeared as a singlet at d 3.68 ppm, and the methyl protons of the Boc-group, which appeared as a singlet at d 1.42 ppm, whereas the methyl protons of the -CHCH 3 moiety yielded a doublet at d 1.27 (J = 7.1 Hz) ppm. In the 13  The crystals of (S)-3b are fully enantiomorphous to the crystals of (R)-3b (Fig. 2). This means that the crystals of (S)-3b and (R)-3b are related, like the le hand is to the right hand. For such crystals, the crystal structures are identical (the same lattice symmetry, equal cell parameters, etc.), except that the molecules of (S)-3b and (R)-3b are enantiomers.
We also tried to analyze the enantiomers (S)-3a-c and (R)-3ac and their unprotected forms by chiral HPLC analysis. Various attempts were made using different enantioselective HPLC columns, but this method proved unsuccessful. Then, we proceeded to determine the enantiomeric purity of our compounds using NMR methods. Many NMR spectroscopic techniques rely on chiral auxiliaries such as chiral derivatization agents, chiral lanthanide shi reagents, metal complexes, and chiral solvating agents. 37,38 Fuertes et al. developed a simple chiral derivatization protocol for the enantiopure determination of chiral primary amines using 1 H NMR spectroscopic analysis. The method involves the condensation of amines with 2-formylphenylboronic acid (2-FPBA) and (S)-1,1 ′ -bi-2-naphthol ((S)-BINOL). This method allows a mixture of diastereomeric derivatives to be obtained, the ratio of which can be determined by integrating the resonances in their 1 H NMR spectra, which makes it easy to determine the enantiopurity of the starting amine. 39 In our case, the synthetic strategy to determine enantiomeric excess (ee) for amines containing remote stereogenic centers is based on the formation of iminoboronate ester complexes (Scheme 2). Deprotection of the N-Boc group from compounds 3a-c was carried out in the presence of TFA, followed by base workup using Cs 2 CO 3 in order to generate free primary amines 4a-c. Furthermore, the reaction of chiral primary amines (S)-4ac and (R)-4a-c with 2-FPBA and stereodened (R)-BINOL in CDCl 3 with 4 Å molecular sieves for 18 h at room temperature afforded a mixture of diastereomeric iminoboronate ester complexes (S,R)-5a-c and (R,R)-6a-c. The diastereomeric ratio (dr) was determined by comparing the integration ratios of distinct protons in their 1 H NMR spectra, thus allowing indirect determination of the enantiopurity of their parent amines 4a-c ( Table 1).
Analysis of the 1 H NMR spectra of the iminoboronate ester complexes (S,R)-5b and (R,R)-5b revealed a characteristic resonance of the methyl protons of the esteric group (COOCH 3 ), which appeared in their 1 H NMR spectra as a singlet at d 3.56 ppm and d 3.54 ppm (chemical shi difference, Dd, between methyl protons of COOCH 3 is 0.02 ppm), respectively ( Fig. 4). NMR analysis showed that the dr of the iminoboronate ester complex (S,R)-5b was 96 : 4, whereas the dr of (R,R)-5b was 94 : 6. Furthermore, this diastereomeric ratio is expected to be in quantitative agreement with the enantiomeric ratio of chiral amines (S)-4b and (R)-4b. Thus allowing us to conclude that their ee are 92% and 88%, respectively. Analysis of the 1 H NMR spectra of each derivatization reaction revealed the presence of at least one pair of resolved diastereomeric resonances in each case, whose integrals could be used to determine indirectly the enantiopurity of their parent amine 4. The 1 H NMR spectrum of the pair of diastereomers (S,R)-5c and (R,R)-5c showed characteristic methyl proton resonances of the ester group (COOCH 3 ) at d 3.57 ppm and d 3.56 ppm, respectively. In this case, the products (S,R)-5c and (R,R)-5c were obtained with 93 : 7 dr (86% ee for (S)-4c) and 100 : 0 dr (100% ee for (R)-4c), respectively. However, the 1 H NMR spectra of the corresponding products, (S,R)-5a and (R,R)-5a, showed that the methyl ester group protons overlapped and resonated at d 3.62 ppm. Therefore, the determination of diastereomers (S,R)-5a and (R,R)-5a according to the diastereomeric ratio by integration of their 1 H NMR spectra showed the distinct resonances of the proton from the -CHCH 3 moiety. The -CHCH 3 fragment gave quadruplets at d 3.21 (J = 7.0 Hz) and d 3.18 (J = 7.0 Hz) ppm, respectively. The investigation of synthesized iminoboronate ester complexes (S,R)-5a and (R,R)-5a produced 1 H NMR spectra with a diastereomeric ratio of 100 : 0 (100% ee for (S)-4a and (R)-4a) for both complexes.
In addition, deprotection of the N-Boc group from compounds 3a-c potentially had no inuence on changing the enantiomeric ratio for obtained amines 4a-c. Therefore, the synthesis of enantiomers 3a-c (Scheme 1), as described above, is highly enantioselective with no or limited epimerization.
The prepared enantiomers, (S)-3a and (R)-3a, are potential synthons in the synthesis of small peptides. Synthetic small peptides, including heterocyclic dipeptides, are thus attractive agents and targets for therapies and diagnostics. [40][41][42] For example, Pavadai et al. reported synthesis of a piperidin-4-one derivative containing dipeptide as an acetyl cholinesterase and b-secretase inhibitor; 43 Blaszczyk et al. synthesized new piperidine dipeptides exhibiting arginase inhibition, with high intracellular activity that could be of use in the treatment of cancer. 44,45 In the present work, we prepared 2-(Boc-amino)piperidine dipeptides (S,S)-7 and (R,S)-7 by coupling enantiomers (S)-3a and (R)-3a with L-phenylalanine (Scheme 3). First, the ester (S)-3a was hydrolyzed with 2 N NaOH in methanol to afford acid (S)-6 in a yield of 90%. Aer that, the reaction of 2- hexa-uorophosphate (HATU) was carried out in the presence of DIPEA in a polar aprotic solvent, such as DMF, at room temperature to form the corresponding active ester. HATU has proven to be a highly reactive peptide coupling reagent, free from by-product formation and product racemization compared to other commonly used coupling reagents. 46 In our case, the corresponding active ester was coupled with Lphenylalanine ethyl ester hydrochloride to produce N-Bocdipeptide (S,S)-7 in a 59% yield. The same method was used to synthesize N-Boc-dipeptide (R,S)-7 from acid (R)-6 with Lphenylalanine. The formation of N-Boc-dipeptides (S,S)-7 and (R,S)-7 was established by NMR analysis. The reaction produced (S,S)-7 in a diastereomeric ratio of 94 : 6 and (R,S)-7 in a diastereomeric ratio of 90 : 10 (ESI in Fig. S38-S43 †). The 1 H NMR spectrum of (S,S)-7 revealed a quadruplet of -CHCH 3 proton at Table 1 Determination of enantiopurity for compounds (S,R)-5a-c, (R,R)-5a-c and 4a-c in 1    We also investigated the transformation of N-Boc-dipeptides (S,S)-7 and (R,S)-7 to N-nosyl-dipeptides (S,S)-9 and (R,S)-9 (Scheme 4). The use of a p-nitrobenzenesulfonyl (nosyl) group to protect the amino functional group is of great importance for obtaining N-methylated amino acids and peptides. 47 Moreover, sulfonamides play a signicant role in medicine as antibiotics, antithyroid agents, and antitumor drugs. 48,49 For instance, Murthy et al. reported a series of novel benzhydryl piperazinecoupled nitrobenzenesulfonamide hybrids as agents which showed excellent anti-tuberculosis activity, 50 and Ugwuja et al. developed and synthesized new peptide-derived antimalaria and antimicrobial agents bearing a sulfonamide moiety. 51 To remove the protecting Boc-group, 52 dipeptide (S,S)-7 was dissolved in DCM and then TFA was added under stirring at room temperature for 30 min. Aer the removal of the solvent in vacuo, the corresponding triuoroacetate (S,S)-8 was used directly in the next step without further purication. The deblocked product, (S,S)-8, was coupled with 4-nitrobenzenesulfonyl chloride in the presence of 1 M Na 2 CO 3 in acetonitrile to obtain the corresponding diastereomeric Nnosyl-dipeptide (S,S)-9 in a yield of 86%. The same reaction conditions as above were applied to synthesize N-nosyldipeptide (R,S)-9 in a yield of 89%. 1  We then performed a nucleophilic substitution reaction with triate esters (R)-2a-c and (S)-2a-c, chiral 3-Bocaminopiperidine and 3-Boc-aminopyrrolidine, to obtain diastereomers 10a-c and 11a-c, respectively (Scheme 5). Optimization of the nucleophilic substitution conditions was undertaken for determination of diastereomeric selectivity, choosing triate esters (R)-2b and (S)-2b as enantiomeric pair. Then, nucleophilic substitution with (R)-3-Boc-aminopiperidine was carried out at different temperatures, such as room temperature, −30°C, and −50°C, and the 1 H NMR spectral data of the crude samples of products (2S,3R)-10b and (2R,3R)-10b were analyzed ( Table 2). The 1 H NMR spectra of (2S,3R)-10b and (2R,3R)-10b revealed characteristic resonance for the doublet signal of the proton of -CHCH(CH 3 ) 2 at d 2.67 (J = 10.8 Hz) ppm and d 2.71 (J = 10.8 Hz) ppm, respectively. In our study, the poorest stereoselectivity was observed when the reaction mixture was stirred at room temperature − 75 : 25 dr for (2S,3R)-10b and 78 : 22 dr for (2R,3R)-10b. Furthermore, when the reaction was carried out at −30°C, the resulting diastereomeric ratios were 87 : 13 dr and 93 : 7 dr, respectively, for (2S,3R)-10b and (2R,3R)-10b. Moreover, carrying out the reaction at −50°C yielded a high stereoselectivity (for (2S,3R)-10b, it was 94 : 6 dr, and for (2R,3R)-10b, it was 100 : 0 dr) and a good yield (for (2S,3R)-10b, yield was 86%, and for (2R,3R)-10b yield was 83%).

Synthesis of triates (2a-c)
A solution of the corresponding ester (1a-c) (1 equiv.) and pyridine (1.2 equiv.) in DCM (0.1 M) was cooled to 0°C and stirred for 5 min under an argon atmosphere. Then, tri-uoromethanesulfonic anhydride (1.2 equiv.) was added dropwise, and the reaction mixture was stirred for 2 hours. The resulting solution was quenched with water (10 mL), and the aqueous layer was separated and extracted with DCM (2 × 15 mL) and brine (15 mL). The organic layer was dried with anhydrous sodium sulfate, ltered, and then concentrated under reduced pressure. Crude product (2a-c) was directly used in the next step without further purication.

Synthesis of alkanoates (3a-c, 10a-c and 11a-c)
Method A. Triate (2a) (500 mg, 1 equiv.) was added to a mixture of N-Boc-cycloamine (1 equiv.) and TEA (1 equiv.) in DCM (15 mL) under an argon atmosphere at −50°C, and the solution was stirred at this temperature for 4 hours. The reaction mixture was diluted with DCM (10 mL) and washed with H 2 O (2 × 15 mL) and brine (15 mL). The organic layer was dried with anhydrous sodium sulfate, ltered, and then concentrated under reduced pressure. The crude product was puried by ash chromatography.
Method B. Triate (2b-c) (500 mg, 1 equiv.) was added to a mixture of N-Boc-cycloamine (1.5 equiv.) and TEA (1.5 equiv.) in DCM (15 mL) under an argon atmosphere at −50°C, and the solution was stirred at r.t. overnight. The reaction mixture was diluted with DCM (10 mL) and washed with H 2 O (2 × 15 mL) and brine (15 mL). The organic layer was dried with anhydrous sodium sulfate, ltered, and then concentrated under reduced pressure. The crude product was puried by ash chromatography.